The present invention relates to an optical detection device, an optical detection method, and a program, and in particular, relates to an optical detection device, an optical detection method, and a program employing Raman spectroscopy technology in the analysis of substances. Namely, the present invention is related to an optical detection device in which two or more beams of pulsed laser light are radiated onto a sample, and substances in the sample are analyzed by observing Raman scattered light that is emitted from the sample as a result.
The detection of trace substance using Raman spectroscopy technology is of great importance as a fundamental technology in analytical devices, and there have been many technological developments therein. With recent advances in medical technology, attempts have been made to apply such trace substance detection in medical diagnostic technology, such that there is now also a demand to further increase detection sensitivity for trace substances in the field of medical diagnostic technology.
The coherent anti-Stokes Raman scattering (CARS) method is known as an example of the Raman spectroscopy technology above (Patent Document 1). This is a method in which two or more types of optical pulse are radiated onto a sample, and then coherent anti-Stokes Raman scattered light (CARS light), which has been emitted from the sample due to a non-linear optical process caused by interaction between the optical pulses, is observed.
For example, consider the use of CARS to observe a molecule having energy levels like those illustrated in FIG. 10(a). First, a molecule of the sample at an initial state energy level L1 is excited by the incidence of a first pulsed light (excitation light) having an angular frequency of ω1, and the energy level of the molecule increases to L3 as indicated by the arrow A. Then, due to causing a second pulsed light (Stokes light) having an angular frequency of ω2 to be incident to the molecule, the energy level of the molecule falls from L3 to L2 as indicated by the arrow B due to photoemission. Then, due to causing a third pulsed light (probe light) having an angular frequency of ω3 to also be incident to the molecule, the energy level of the molecule rises from L2 to L4 as indicated by the arrow C, and then falls from L4 to L1 as indicated by the arrow D due to emitting CARS light.
Thus, what is known as a four wave mixing process occurs due to the incidence of the three types of pulsed light, having angular frequencies of ω1, ω2, and ω3, and CARS light having an angular frequency of ω1+ω3−ω2 is emitted as a result. CARS light of this sort arises particularly intensely when the frequency difference between the incident pulses, given by Δω=ω1−ω2, resonates with the difference in the energy levels of the molecule being observed. When pulsed light that can actually be used in practice is considered, then considering that a strong signal is obtained when Δω matches a vibrational mode frequency of the molecule, molecules having such vibrational modes can be detected. This method can also be implemented using two types of pulsed light, so as to achieve the detection of CARS light having an angular frequency of 2ω1−ω2 by using the first pulsed light to induce the optical process that would have been caused by the third pulsed light.
FIG. 10(b) illustrates a spectrum SP of pulsed light radiated onto the sample, and a spectrum SC of CARS light emitted as a result of the radiation. Pulsed light corresponding to a portion of the spectrum SP gives rise to Raman scattering, and the spectrum SC of CARS light is emitted with the position of wavelength λ shifted by Δλ toward the short wavelength side. The width Δλ of the wavelength shift is generally known as the Raman shift, and is sometimes expressed in wavenumbers n (the reciprocal of wavelength λ, cm−1) instead of wavelength. References in the following to the wavelength of pulsed light indicate the central wavelength of the spectrum of pulsed light.
FIG. 11 illustrates a Raman spectroscopy device 80 according to related technology that uses these fundamental principles. The Raman spectroscopy device 80 is configured including two types of laser pulse light sources, these being a first laser pulse light source 82 and a second laser pulse light source 84, an optical system 86 for radiating pulsed light from these light sources onto a single location of a sample 88, and a detection device 90 that detects CARS light emitted by the sample 88 (Patent Document 1, and Non-patent Document 1). Then, for example, CARS light emitted by a specific molecule included in the sample 88 can be selectively detected by changing the wavelengths of the pulsed light emitted from the first laser pulse light source 82 and from the second laser pulse light source 84.
Since CARS light is detected due to molecular vibration, which is an intrinsic property of the molecule in the sample 88, there is no need to stain a trace quantity of a molecule using a marking substance or the like during, for example, the identifying of a trace quantity of a molecule in a living organism. Accordingly, observations can be made without being hindered by the influence of a marking substance, particularly when observing a small molecular compound formed from molecules that are smaller than the molecules of the marking substance. Thus, Raman spectroscopy devices based on the observation of CARS light are particularly advantageous over those employing other methods when observing living organism.